韩国延世大学医学院Hyongbum Henry Kim团队，使用先导编辑对耐药基因变异进行饱和分析。2024年11月12日，《自然—生物技术》杂志在线发表了这项成果。

研究人员介绍了PEER-seq，这是一种基于先导编辑的高通量方法，可评估单核苷酸变异（SNV）的功能效应。PEER-seq使用先导编辑引入预定的SNV和同义标记突变，并通过深度测序内源性靶向区域，识别引入的SNV。

研究人员在表皮生长因子受体基因（EGFR）中生成并功能性评估了2476个SNV，涵盖了典型酪氨酸激酶结构域中99%的所有可能变异。

研究人员在PC-9细胞中测定了在共存替代突变T790M存在的情况下，95%所有可能的EGFR蛋白变异体，对常见酪氨酸激酶抑制剂（阿法替尼、奥希替尼和厄洛替尼）的耐药性谱。该研究有潜力显著提高临床治疗选择的精准度。

据了解，用于表征不确定意义基因变异（VUS）功能效应的方法，因突变空间的覆盖不完全而受到限制。在临床肿瘤学中，VUS引起的药物耐药性可能妨碍最佳治疗的实施。

附：英文原文

Title: Saturation profiling of drug-resistant genetic variants using prime editing

Author: Kim, Younggwang, Oh, Hyeong-Cheol, Lee, Seungho, Kim, Hyongbum Henry

Issue&Volume: 2024-11-12

Abstract: Methods to characterize the functional effects of genetic variants of uncertain significance (VUSs) have been limited by incomplete coverage of the mutational space. In clinical oncology, drug resistance arising from VUSs can prevent optimal treatment. Here we introduce PEER-seq, a high-throughput method based on prime editing that can evaluate the functional effects of single-nucleotide variants (SNVs). PEER-seq introduces both intended SNVs and synonymous marker mutations using prime editing and deep sequences the endogenous target regions to identify the introduced SNVs. We generate and functionally evaluate 2,476 SNVs in the epidermal growth factor receptor gene (EGFR), including 99% of all possible variants in the canonical tyrosine kinase domain. We determined resistance profiles of 95% of all possible EGFR protein variants encoded in the whole tyrosine kinase domain against the common tyrosine kinase inhibitors afatinib, osimertinib and osimertinib in the presence of the co-occurring substitution T790M, in PC-9 cells. Our study has the potential to substantially improve the precision of therapeutic choices in clinical settings.

DOI: 10.1038/s41587-024-02465-z

Source: https://www.nature.com/articles/s41587-024-02465-z